Eosinophils have functions in health and in the pathogeneses of asthma, allergies and other respiratory tract diseases. As cells of the innate immune system, a distinct attribute of eosinophils is their content of multiple already synthesized cytokines stored within eosinophil cytoplasmic granules and vesicles along with other granule cationic proteins. Eosinophil functions are based on their regulated release of diverse granule-derived cytokines and proteins that mediate interactions with other cells in the microenvironment of tissue sites. Defining functional interactions between eosinophils, mediated by their secreted cationic and cytokine proteins, and other cells in tissues is germane to understanding eosinophil functions in both ongoing immune homeostasis and acute and chronic diseases. Eosinophils in tissues, including airways and lung sites, exhibit ultrastructural evidence of two forms of degranulation: 1) piecemeal degranulation by which granule contents are mobilized into and secreted by cytoplasmic secretory vesicles, and 2) release of intact granules extruded by cytolysis from within eosinophils. We have established that cell-free eosinophil granules express ligand- binding cytokine, chemokine and eicosanoid receptors that activate intragranular signaling to stimulate granules to directly secrete selected granule-derived proteins. Both agonist-specific piecemeal degranulation from within intact eosinophils and secretion from within free granules can release, selectively and differentially, specific granule cationic proteins and some of the multitude of cytokines stored preformed within granules. Mechanisms regulating selective release of eosinophil granule-derived proteins remain to be delineated. Studies will investigate mechanisms involved in the regulated release of eosinophil-derived proteins, including cytokines and cationic proteins. The overall hypothesis is that, central to the functions of eosinophils, the intracellular compartmentalization, trafficking and release of relevant eosinophil proteins from eosinophils and their free granules are dependent on regulatory mechanisms that with stimulus-dependent specificity can finely regulate the differential and selective secretion of cytokines. Aims are to investigate: 1) mechanisms regulating the secretion competence of cell-free eosinophil granules; 2) mechanisms mediating the secretion of cytokines and other proteins from within eosinophils; and 3) stimulus-dependent mobilization and targeted release of eosinophil cytokines. Understanding molecular and cellular mechanisms that regulate the extracellular release of eosinophil granule-derived proteins will provide insights into eosinophil functions that involve interactions between eosinophils and other cells in tissue sites in the lung and elsewhere. PUBLIC HEALTH RELEVANCE: Eosinophil-associated diseases, including asthma, allergies and other disorders, are increasingly prevalent diseases and major public health problems. Better understanding of the functions of eosinophils in these diseases may lead to improved therapies for these diseases.